DNA replication and DNA repair in Acholeplasma laidlawii would be studied with the following specific aims: Folded chromosomes would be isolated and their properties as related to the DNA replication cycle examined using sedimentation and electron microscopy methods. Folded chromosomes would be used to study the interaction of the chromosome with the cell membrane. Coliphages M13, phix174, and G4 DNA and folded chromosomes would be used as substrates in in vitro DNA synthesis studies comparing A. laidlawii and E. coli protein extracts. DNA replication proteins from A. laidlawii would be isolated and studied. In in vivo studies, Okazaki fragments and RNA primers would be sought. The initiation process would be studied using amino acid starvation and antibiotic inhibition of reinitiation. Isolation of restriction fragments containing the orgin would be done using restriction fragment analysis of cells labeled at the orgin. Orgin-RNA would be sought in the origin restriction fragments. Attempts would be made t isolate temperature sensitive dna mutants of A. laidlawii. Excision repair and inducible post-replication DNA repair systems would be sought and characterized. Restriction-modification systems, A. laidlawii plasmids, and insertion sequences would be investigated. The primary cellular target for transformation of a cell into a malignant cell is the cellullr DNA. Detailed knowledge of DNA replication and DNA repair processes are critically important in understanding this transformation process. Most Mycoplasmatales are parasitic, and some are pathogenic for important domestic animals and for man, demonstrating their economic and medical importance. Thus, this project would study critically important processes to the cancer problem, in organisms of direct medical and economic importance, demonstrating its direct relevance to the health sciences.